Authors
Muhammad N Toor, Abdus Samad, Yuliang Huang, Xianliang Huang, Xiao Han, Wei Zhou, Xiangrui Liu, Lei Luo, Hao Shao
Published in
European journal of medicinal chemistry. Volume 321. Issue Pt 1. Pages 119393. Oct 01, 2026. Epub Oct 01, 2026.
Abstract
DNA polymerase theta (Polθ) is an emerging therapeutic target in homologous recombination-deficient cancer, yet fluorescent probes for Polθ inhibitor screening and cellular imaging remain largely unexplored. To address this gap, we report the rational design, synthesis, and biological evaluation of small-molecule fluorescent probes targeting the polymerase domain of Polθ (Polθ-pol). These probes were synthesized by conjugating a Polθ-pol recognition scaffold to fluorescein- and 4-chloro-7-nitrobenzofurazan-derived fluorophores through flexible linkers. Spectroscopic characterization revealed solvent-dependent absorption and emission profiles. Fluorescence polarization assay demonstrated concentration-dependent and saturable binding of all probes to Polθ-pol, with Kd values in nanomolar range. Among the series, Fluorescein-derived Probe-2 exhibited the strongest binding affinity and enabled a robust FP competition assay for successful evaluation of Polθ inhibitors, establishing it as a promising fluorescent tracer for inhibitor screening. Furthermore, live-cell fluorescence imaging showed that NBD-derived probes displayed comparatively stronger cellular fluorescence. Probe-5 showed co-localization with Polθ and increased γ-H2AX foci formation in HCT-116 cells. Collectively, this study provides the first set of small-molecule fluorescent probes for both Polθ-pol inhibitor screening and investigation of Polθ-associated processes in living cells.
PMID:
42828964
Bibliographic data and abstract were imported from PubMed on 04 Oct 2026.
Read full publication at:
Please sign in
to see all details.
Advertisement
Stats
- Recommendations n/a n/a positive of 0 vote(s)
- Views 12
- Comments 0